Esters of protoverine



United States Patent 3,066,142 ESTERS OF PROTOVERINE S. Morris Kupchan,Madison, Wis., assignor to Wisconsin Alumni Research Foundation,Madison, Wis., a corporation of Wisconsin No Drawing. Filed July 11,1960, Ser. No. 41,793 7 Claims. (Cl. 260-287) The present inventionrelates to novel ester derivatives of protoverine.

The structures of the naturally occurring hypotensive protoverineesters, prot-overatrine A and protoveratrine B have recently beenelucidated. Kupchan, S. Morris et al., I. Am. Chem. Soc. 81, 1009(1959); 82, 2616 (1960). These two alkaloids have been used to someextent in recent years for treatment of hypertension but the deterrentto wider use is the narrow range between their therapeutic and emeticdoses. L. S. Goodman and A. Gilman "The Pharmacological Basis ofTherapeutics, The Mac- Millan Co., New York, N. Y., 2nd edition, pp.747-754 (1955); O. Krayer and V. A. Drill, Pharmacology in GENERALPROCEDURES Acylations with limited amounts of isobutyryl chloride oracetyl chlride.--To the dry starting material in reagent grade pyridine(l g. in ca. ml. pyridine) cooled in an ice bath, was added slowly thespecified quantity of acyl chloride. The flask was protected frommoisture with a calcium chloride tube-and placed in an ice-water bathwhich was allowed to warm gradually to room temperature. After -30hours, the solution was transferred to a separatory funnel and treatedwith chloroform, ice-water, and dilute ammonium hydroxide to pH 8-9. Thesolution was extracted with chloroform 4 times; the combined extractswere dried over anhydrous sodium sulfate and evaporated to dryness underreduced pressure. To remove all traces of pyridine, the residue wasrepeatedly. dissolved in benzene and evaporated to dryness.

Methanolyses of 16-acetate 'esters.-The ester (5 to 10 mg.) wasdissolved in methanol (0.1 to 0.3 ml.) and left at room temperature in acorked flask. The course of the reaction was followed by paperchromatography using the solvent systems of Levine, I. et al., J. Am.Pharm. Assoc, 44, 543 (1955), and the methanolysis was interrupted whenthe mixture appeared to be most suitable for a column separation. Thepreparative-scale reaction was then run in the corresponding manner.

Column separations on alumina.-The partial acylations andrrrethanoly-ses afforded mixtures of products. In all cases, separationswere effected by chromatography on acid-washed alumina (20-25 g. per g.of alkaloid mixture). The solvent mixtures were selected on the basis ofthe R -values of the alkaloids, and generally ranged from benzene,benzene-chloroform, and chloroform, to mixtures of chloroform-methanolcontaining gradually increasing proportions of methanol. The acylationswere usually accompanied by some discoloration. The colored material wasgenerally small in quantity and the major proportion was either retainedby the alumina or eluted with the first few fractions. Initial fractionscollected were usually kept very small in order to separate the yellowor brown impurities in the forerun from easilyeluted colorlessalkaloids.

Recrystallizati0ns.-Most of the compounds obtained evaporation of thesolutions eluted from the columns contained slight residual amounts ofyellow to brown contaminants. The colored material was separated fromeach resin by dissolution in a little ether or acetone and dropwiseaddition of petroleum ether to turbidity filtration at this pointremoved colored impurity and gave a colorless filtrate from whichcrystalline product separated on standing.

Melting points are corrected for stem exposure. Values of [oc] have beenapproximated to the nearest degree. Ultraviolet absorption spectra weredetermined in ethanol on a Cary recording spectrophotometer (model 11MS). Infrared spectra were determined in chlorofform on a Baird doublebeam recording spectrophotometer.

EXAMPLE 1 Protoverine 6,15-diisobutyrate (II) and protoverin3,6,I5-triisobutyrate (III).These esters were obtained as the principalproducts of acrylation of protover'ine with 2.3 mole equivalents ofisobutyryl chloride for 17 hours. The products were separated asdescribed above. No indication of more highly acylated products wasnoted.

EXAMPLE 2 Protoverine 3,7,16-triacetate 6,15-diisobutyrate (IV)l-I,Protoverine 6,15-diisobutyrate (II) (500 mg.) was treated with pyridine(3 ml.) and acetic anhydride (10 ml.) and the mixture was heated for 2.5hours in a water bath at 80 C. Excess acetic anhydride was cautiouslydecor n; posed by dropwise addition of methanol (7 ml.) and the reactionmixture was worked up in the usual manner ice-water, chloroform andammonia. The residue ob; tained by evaporation ofthe chloroform solutionto dryness was crystallized from acetone-petroleum ether in the form ofcolorless needles.

EXAMPLE 3 Protoverine 3,6,15-triis0butyrate 7,16-diacetate V)Acetylation of III by the procedure described for preparation of IVafforded the crystalline diacetate fromacetonepetroleum ether.

EXAMPLE 4.

Protoveriwe 3,6,16-triacetate 7,15-diisobutyrate (X) and protoverine3,6,l6-triacetate 15-isobutyrwte (IX) .Protoverine 3,6,l6-triacetate(3.1 g.) in pyridine ml.), I. Am. Chem. Soc., 82, 2242 (1960), wasacylated with isobutyryl chloride (2.0 ml., ca. 4 mole equivalents).

Workup and column separation in the usual manner yielded X as needlesfrom acetone-petroleum ether, and IX as prisms from acetone-petroleumether.

EXAMPLE 7 Proioverine 3,6,7,16-te'traacetate 15-isobutyrate (XI)Aeetylat-ion of IX (500 mg.) by the procedure described for preparationof IV yielded crystalline XI from chloroform-petroleum ether; 7 V iEXAMPLE 8 Protoverine 3,6-diacetate 7,15-diisbutyrate (XII).Methanolysis of X (700 mg.) followed by column separation yielded XIIasprisms from acetone-petroleum ether.

EXAMPLE 9 Protoverin 3,6,7-triacetate -z's0butyrate (XIII). Methanolysisof XI (300 mg.) followed by column separation yielded XIII, crystallizedfrom acetone-petroleum ether.

EXAMPLE 10 EXAMPLE 11 Protoverine 3,7,l5-triacetate 6-is0butyrate (XI111).- Protoverine 3,7,15,16-tetraacetate 6-isobutyrate XIX (1.0 g.) inacetone ml.) and methanol (40 ml.) was allowed to stand at roomtemperature for 60 hours. Column separation yielded XVIII as prisms fromacetonepetroleum ether. EXAMPLE 12 Protoverine 3,6-diis0butyrate (XX ).Asolution of protoverine 14,15-acetonide 3,6-diisobutyrate (1.0 g.), J.Am. Chem. Soc., 82, 2242 (1960), in 2% hydrochloric acid (50 ml.) wasallowed to stand at room temperature for 17 hours. The solution was madealkaline with ammonium hydroxide and extracted exhaustively withchloroform. The chloroform extract was dried over anhydrous sodiumsulfate and evaporated to dryness under reduced pressure. The residue(XX) was crystallized from ether and recrystallized fromacetone-petroleum ether.

' EXAMPLE 13 Protoverine 3,6 diisobutyrate 7,15,16 triacetate(XXl).Acetylation of XX (1.0 g.) by the procedure described for thepreparation of IV yielded XXI, crystallized from ether.

' EXAMPLE 14 Protoverine 3,6-diis0butyrate 7,15-diacetate (XXIIMethanolysis of XXI (1.1 g.) by the procedure described for preparationof XVIII yielded XXII as prisms from acetone-petroleum ether.

EXAMPLE 1 5 EXAMPLE 1? Protoverir ze 3,15,16-triacezate 6-isobutyrate(XXIV).- Protoverine 3,16-diacetate 6-isebutyrate (XXV) (1.8 g.) inpyridine (40 ml.) was acetylated with acetyl chloride (0.3 ml., ca. 1.7mole equivalents). Workup and column separation in the usual manner gaveXXIV, crystallized from ether.

EXAMPLE 1s Protoverine 3,61 diacetate 6,7,]5 triisobutyrate (XXVI).Protoverine 3,16 diacetate 6 isobutyrate (XXV) (1.55 g.) in pyridine(50 ml.) was treated with 4 isobutyryl chloride (1.6 ml., ca. 7 moleequivalents). Workup and column separation in the usual manner yieldedchromatographically homogeneous XXVI. Crystallization from ether yieldedcolorless prisms.

EXAMPLE 19 Protoverine 3,15,16 triacerate 6,7 diz'sobutyrate(XXVH).Protoverine 3,15,16-triacetate 6-isobutyrate (XXIV) (600 mg.) inpyridine (30 ml.) was treated with isobutyryl chloride (0.8 cc., ca. 9mole equivalents). Workup and column fractionation in the usual mannergave XXVII which resisted all attempts at crystallization.

EXAMPLE 2O Protoverine 3,15 diacetate 6,7 diisobutyrate (XX VIII).Methanolysis of XXVII (210 mg.) by the procedure described forpreparation XIII yielded mg. of chromatographically pure XXVIII.

EXAMPLE 21 Protoverine 3,acetate 6,7,15-triisobutyrate (XXIX).-Methanolysis of XXVI (650 mg.) followed by column separation yieldedchromatographically homogeneous amorphous XXIX.

EXAMPLE 22 Protoverine 3,6-a'iis0butyrate 15-acetate (XXX Protoverine3,6-diisobutyrate (XX) (1.1 g.) in pyridine (40 ml.) was treated withacetyl chloride (0.3 cc., ca. 2.5 mole equivalents). Workup and columnseparation in the usual manner yielded XXX as needles from ether.

EXAMPLE 23 Protoverine 3,6,7-triis0butyrate J5-acetate (XXXl).-Protoverine 3,6-diisobutyrate IS-acetate (XXX) (300 mg.) in pyridine (10ml.) was treated with isobutyryl chloride (0.3 ml., ca. 7 moleequivalents). Workup and column separation in the usual manner yieldedXXXI as prisms from ether.

EXAMPLE 2% Protoverine 3,6,15,16 tetraacetate 7 isobutyrate(XXXIII).-Protoverine 3,6,15,16 tetraacetate (850 mg.), J. Am. Chem.Soc, 82, 2616 (1960), in pyridine 10 ml.) was treated with isobutyrylchloride (0.9 ml., ca. 7.5 mole equavilanets) and the solution was keptat room temperature for 12 hours and then at 60 C. for 6 hours. Workupin the usual manner, followed by two column separations yielded XXXIII.

EXA MPLE 25 Protoverine 3,6,15 triacetate 7 isobutyrate(XXXIV).Methanolysis of XXXIII mg.) by the procedure used for thepreparation of XIII yielded XXXIV as prisms from ether.

EXAMPLE 2e Protoverine 3,7,15 triisobutyrate 6,16 diacetate (XXXVII) andprotoverine 3,15,-diis0butyrate 6,16-diacetate (XXX VI ).--Protoverine6,16-diacetate (5.3 g.), I. Am. Chem. Soc., 82, 2616 (1960) in pyridine(60 ml.) was treated with isobutyryl chloride (2.7 ml., 3 moleequivalents). Workup and column separation in the usual manner yieldedXXXVII and XXXVI. Crystallization of XXXVII from acetone-petroleum etheryielded needles.

EXAMPLE 27 Protoverine 3,15 diisobutyrate 6,7,16 zriacetatc(XXXVIII).--Acetylation of XXXVI (900 mg.) by the procedure used forpreapration of IV yielded XXXVIII as prisms from acetone-petroleumether.

EXAMPLE 28 Protoverine 3,7,15 triisobutyrale 6 acetate(XXXIX).-Methanolysis of XXXVII (1.4 g.) followed by column separationyielded chromatographicallyhomogeneous amorphous XXXIX.

EXAMPLE 29 Protoverine 3,15 diisobuzyrare 6,7 diacetate (XL).-Methanolysis of XXXVIII (300 mg.) for 40 hours, followed by columnseparation yielded XL as prisms from ether.

EXAMPLE 3O EXAMPLE 31 Protoverine 3,7 diisobutyl'ate 6,15,16 triacezate(XLII).Protoverine 6,15,16-triacetate (XLI) (500 mg.) in pyridine ml.)was treated with isobutyryl chloride (0.4 ml., ca. 5 mole equivalents).Workup and column separation in the usual manner yielded XLII whichresisted all attempts at crystallization.

EXAMPLE 32 Protoverine 3,7 diisobutyraze 6,15 diacetate(XLlI1).Methanolysis of XLII (300 mg.) followed by column separationyielded XLIII as prisms from ether.

EXAMPLE 33 Protoverine 6,15-diacetale (XLIV) and protoverine 6- acetate(XLVII).Protoverine (6.3 g.) in pyridine (75 ml.) was treated withacetyl chloride (1.6 ml., ca. 1.8 mole equivalents). Workup and columnseparation in the usual manner gave XLIV, crystallized fromchloroform-petroleum ether and XLVII, crystallized fromacetone-petroleum ether.

EXAMPLE 3' 1.

Protoverine 3-is0butyrate 6,15-diacetate (XLV).- Protoverine6,15-diacetate (XLIV) (1.2 g.) in pyridine (50 ml.) was treated withisobutyryl chloride (1.2 ml., 4 mole equivalents). Workup and columnseparation in the usual manner yielded chromatographically-homogeneousamorphous XLV.

EXAMPLE 35 Protoverine 3-isobutyrate 6,7,15-triacetate (XLVI).-Protoverine 3-isobutyrate 6,15-diacetate (XLV) (600 mg.) in pyridine (4ml.) and treated with acetic anhydride (0.17 ml., ca. 2 moleequivalents). After hours at room temperature, Workup and columnseparation in the usual manner gave XLVI as prisms from acetonepetroleumether.

EXAMPLE 36 Protoverine 3,]5-diis0butyrate 6-acetate (XLVIII).-Protoverine 6-acetate (XLVII) (1.7 g.) in pyridine ml.) was treated withisobutyryl chloride (1.6 ml., ca. 5 mole equivalents). Workup and columnseparation in the usual manner yielded chrornatographically-homogeneousamorphous XLVIII.

EXAMPLE 37 Acetylation of protoverine 3,15-diis0butyrate 6-acetate(XLVIII) to protoverine 3,15-diis0butyrate 6,7-diacetate(XL).-Protoverine 3,15-diisobutyrate 6-acetate (XLVIII) (750 mg.) inpyridine (20 ml.) was treated with acetyl chloride (0.5 ml., ca. 7 moleequivalents). After 14 hours, workup and column separation yielded XL,MP.

6 221-222 dec., [u] 41 (0, 1.00 pyr.); infrared spectrum in chloroformidentical with that of sample described above in Example 29.

EXAMPLE 38 Protoverine 3,15-diis0butyrate (XLIX).-Methanolysis of XLVIII(250 mg.) for 40 hours yielded XLIX, crystalized from ether.

The following table gives the melting points and [on] data for thecompounds described above.

Table Compound Protoverine Derivative M.P., [04],,

6, lfi-diisobutyrate 190-191 34 3,7,16-triisobutyrate amorph 113,7,16-triacetate 6,15-diisobutyrate 254-255 49 3,6,15-triis0butyrate7,16-diacetate 234-236 -44 3,7,-diacetate 6,15-diisobutyrate 237-238 -463,6,15-triisobutyrate 7-acetate 234-235 46 3,6,16-triaeetate15-is0butyrate 228-229 3 3,6,16-triacetate 7,15-diis0butyrate 224-225-45 3,6,7,16-tetraaeetate lfi-isobutyrate-.- 258-259 --46 3,6-diacetate7,15-diis0butyrate 252-253 46 3,6,7-triacetate 15-isobutyrate 248-249 403,7,15-triaeetate 6-isobutyrate 265-266 41 3,7,15,16-tetraaeetate6isobutyrat 219-220 51 3,6-diisobutyrate -170 -10 3,6-diis0butyrate7,15,16-triaeetate 210-212 -44 3,6-diis0butyrate 7,15-diacetate 262-26340 14,15-acet0nide G-isobntyrate 3,16- 252-253 +21 diacetate.

3,15,16-triaeetate fi-isobutyrate 177-179 4 3,16-diaeetate S-isobutyrate232-233 -7 3,16-diaeetate 6,7,15-triisobutyrat 233-234 393,15,16-triaeetate 6,7-diisobutyrat amorph. 50 3,15-diacetate6,7-diisobutyrate 227-228 -42 3-aeetato 6,7,15-triisooutyrate amorph 383,6-diisobutyrate 15-acetate -16 -7 3,6,7-triisob utyrate iii-acetate239-241 40 3,6,15,16-tetraaeetate 7-isobutyrate 257-259 -463,6,15-triaeetate 7-isobntyrate 243-244 48 3,15-diisobutyrate6,16-diacetate amorph 16 3,7,15-triisobutyrate 6,16-diacetat 232-233 373,15-diisobutyrate 6,7,16-triaeetate.-. 259-260 -483,7,15-triis0butyrate G-acetate amorph. -30 L 3,15-diisobutyrate6,7-diacetate 222-223 -42 XLI 6,15,16-triaeetate 242-243 18 XLII.3,7-diisobutyrate 6,15,16-triacetate--- amorph 40 XLIII3,7-diisobutyrate 6,15-diacetate 214-216 -37 XLIV 6,15-diacetate 235-23627 XLV 3-isobutyrate 6,15-diacetate. amorph 10 XLVI 3-isobntyrate6,7,15-triaeetate 249-250 39 XLVI G-acetate 180-190 -15 XLVIII3,15-diisobutyrate fi-aeetate -12 XLIX 3,15-diisobutyrate -190 10 Theproducts which appear to be of most interest at the present time areesterified in positions 3 and 15 and unesterified in position 16.Although the ester group at the position 3 can be either acetate orisobutyrate, esterification at position 15 with a branched chain acid(isobutyrate group) appears advantageous.

The novel ester products described above are characterized byinsecticidal properties and can be applied in this field in standarddiluents or carriers including dusts and liquids such as kerosene. Theyhave been found effective (LD/SO) against ordinary house flies indilutions as low as 2 mg. per liter of diluent. For most purposes,concentrations of around .01-1.0% by weight are generally recommended.

I claim:

1. Protoverine 3,6,15-triisobutyrate 7-acetate.

2. Protoverine 3,7,15-triisobutyrate 6-acetate.

3. Protoverine 6,7,15-triisobutyrate 3-acetate.

4. Protoverine 3,15-diisobutyrate 6,7-diacetate.

5. Protoverine 6,15-diisobutyrate 3,7-diacetate.

6. Protoverine 7,15-diisobutyrate 3,6-diacetate.

7. Protoverine 3,6,7-triacetate 15-isobutyrate.

References Cited in the file of this patent Kupchan et al.: Chemistryand Industry, December 6, 1958, page 1626.

1. PROTOVERINE 3,6,15-TRIISOBUTYRATE 7-ACETATE. 